Bottom Line:
Lower incisors and molars did not erupt.These results indicate that local activation of β-catenin in the osteoblasts and odontoblasts leads to aberrant dento-alveolar complex formation.Therefore, appropriate inhibition of Wnt/β-catenin signaling is important for the dento-alveolar complex formation.

Affiliation: Cluster for Craniofacial Development and Regeneration Research, Institute of Oral Biosciences, Chonbuk National University School of Dentistry, Jeonju, Korea.

ABSTRACTWnt/β-catenin signaling plays a critical role in bone formation and regeneration. Dentin and cementum share many similarities with bone in their biochemical compositions and biomechanical properties. Whether Wnt/β-catenin signaling is involved in the dento-alveolar complex formation is unknown. To understand the roles of Wnt/β-catenin signaling in the dento-alveolar complex formation, we generated conditional β-catenin activation mice through intercross of Catnb(+/lox(ex3)) mice with Col1a1-cre mice. In mutant mice, tooth formation and eruption was disturbed. Lower incisors and molars did not erupt. Bone formation was increased in the mandible but tooth formation was severely disturbed. Hypomineralized dentin was deposited in the crown but roots of molars were extremely short and distorted. In the odontoblasts of mutant molars, expression of dentin matrix proteins was obviously downregulated following the activation of β-catenin whereas that of mineralization inhibitor was increased. Cementum and periodontal ligament were hypoplastic but periodontal space was narrow due to increased alveolar bone formation. While cementum matrix proteins were decreased, bone matrix proteins were increased in the cementum and alveolar bone of mutant mice. These results indicate that local activation of β-catenin in the osteoblasts and odontoblasts leads to aberrant dento-alveolar complex formation. Therefore, appropriate inhibition of Wnt/β-catenin signaling is important for the dento-alveolar complex formation.

Figure 1: Targeted activation of β-catenin in the dentoalveolar complex. (A) In immunohistochemistry, β-catenin is localized in the Am, Od, and Ob of developing mouse dentoalveolar complex at P8. (B) β-galactosidase activities are also found in the Od and Ob of Col1a1-cre:R26R double transgenic mouse dentoalveolar complex at P8. (C) Col1a1-cre:Catnb+/lox(ex3) mutant mice exhibit severe growth retardation with short height and small body weights. (D) Differences in body weights between WT and MT mice are appeared in 3 weeks after birth and are clearer in the mice at 4 weeks-old. (E) Genotype analysis of mutant mice after intercross with Col1a1-cre and Catnb+/lox(ex3) mice. (F-I) Gross appearance of incisors in WT and MT mice at 4 weeks-old. Both of upper and lower incisors are normally erupted in WT mice, whereas upper incisors are erupted but lower incisors (white arrow) are not erupted in MT mice. (J, K) Molars of lower jaw (white arrowheads) are not erupted into oral cavity in MT mice in contrast to normally erupted in WT mice. Am, ameloblasts; Od, odontoblasts; Ob, osteoblasts; WT, wild type; MT, mutant. *P<0.05. Scale bar=200 µm (A, B).

Mentions:
In the mandible of mouse at P8, β-catenin was localized in the ameloblasts, odontoblasts, and osteoblasts (Fig. 1A). β-galactosidase activities were also observed in the odontoblasts and osteoblasts of the Col1a1-cre:R26R double transgenic mouse at P8 (Fig. 1B). This confirmed that targeting of β-catenin with Col1a1-cre was effective in the formation of dento-alveolar complex. At 4 week-old, height of MT mice was shorter than that of WT mice (Fig. 1C). Body weight was also decreased in the MT mice compared to the WT littermates (Fig. 1D). The mean body weight was 7.77±0.61 g (n=9) and 3.58±1.44 g (n=9) in the WT and MT mice at 3 week-old, respectively. The decrease of body weight was more evident in the 4 week-old MT mice (Fig. 1D). The mean body weight was 13.55±1.42 g (n=9) and 3.73±0.72 g (n=9) in the WT and MT mice at 4 weeks-old, respectively. The decrease of body weight was statistically significant (P<0.05). In genotyping with tail DNA, bands of 720 bps and 600 bps were amplified for cre sequences and deleted allele for β-catenin exon 3 loci (Fig. 1E). In stereoscopic observation of incisors, both of upper and lower incisors were normally erupted in the WT mice. In the MT mice, upper incisors were erupted but lower incisors were not erupted (Fig. 1F-I). Mandibular molars were normally erupted into the oral cavity in WT mice but those of MT mice were not erupted as the same as lower incisors (Fig. 1J, K).

Figure 1: Targeted activation of β-catenin in the dentoalveolar complex. (A) In immunohistochemistry, β-catenin is localized in the Am, Od, and Ob of developing mouse dentoalveolar complex at P8. (B) β-galactosidase activities are also found in the Od and Ob of Col1a1-cre:R26R double transgenic mouse dentoalveolar complex at P8. (C) Col1a1-cre:Catnb+/lox(ex3) mutant mice exhibit severe growth retardation with short height and small body weights. (D) Differences in body weights between WT and MT mice are appeared in 3 weeks after birth and are clearer in the mice at 4 weeks-old. (E) Genotype analysis of mutant mice after intercross with Col1a1-cre and Catnb+/lox(ex3) mice. (F-I) Gross appearance of incisors in WT and MT mice at 4 weeks-old. Both of upper and lower incisors are normally erupted in WT mice, whereas upper incisors are erupted but lower incisors (white arrow) are not erupted in MT mice. (J, K) Molars of lower jaw (white arrowheads) are not erupted into oral cavity in MT mice in contrast to normally erupted in WT mice. Am, ameloblasts; Od, odontoblasts; Ob, osteoblasts; WT, wild type; MT, mutant. *P<0.05. Scale bar=200 µm (A, B).

Mentions:
In the mandible of mouse at P8, β-catenin was localized in the ameloblasts, odontoblasts, and osteoblasts (Fig. 1A). β-galactosidase activities were also observed in the odontoblasts and osteoblasts of the Col1a1-cre:R26R double transgenic mouse at P8 (Fig. 1B). This confirmed that targeting of β-catenin with Col1a1-cre was effective in the formation of dento-alveolar complex. At 4 week-old, height of MT mice was shorter than that of WT mice (Fig. 1C). Body weight was also decreased in the MT mice compared to the WT littermates (Fig. 1D). The mean body weight was 7.77±0.61 g (n=9) and 3.58±1.44 g (n=9) in the WT and MT mice at 3 week-old, respectively. The decrease of body weight was more evident in the 4 week-old MT mice (Fig. 1D). The mean body weight was 13.55±1.42 g (n=9) and 3.73±0.72 g (n=9) in the WT and MT mice at 4 weeks-old, respectively. The decrease of body weight was statistically significant (P<0.05). In genotyping with tail DNA, bands of 720 bps and 600 bps were amplified for cre sequences and deleted allele for β-catenin exon 3 loci (Fig. 1E). In stereoscopic observation of incisors, both of upper and lower incisors were normally erupted in the WT mice. In the MT mice, upper incisors were erupted but lower incisors were not erupted (Fig. 1F-I). Mandibular molars were normally erupted into the oral cavity in WT mice but those of MT mice were not erupted as the same as lower incisors (Fig. 1J, K).

Bottom Line:
Lower incisors and molars did not erupt.These results indicate that local activation of β-catenin in the osteoblasts and odontoblasts leads to aberrant dento-alveolar complex formation.Therefore, appropriate inhibition of Wnt/β-catenin signaling is important for the dento-alveolar complex formation.

Affiliation:
Cluster for Craniofacial Development and Regeneration Research, Institute of Oral Biosciences, Chonbuk National University School of Dentistry, Jeonju, Korea.

ABSTRACTWnt/β-catenin signaling plays a critical role in bone formation and regeneration. Dentin and cementum share many similarities with bone in their biochemical compositions and biomechanical properties. Whether Wnt/β-catenin signaling is involved in the dento-alveolar complex formation is unknown. To understand the roles of Wnt/β-catenin signaling in the dento-alveolar complex formation, we generated conditional β-catenin activation mice through intercross of Catnb(+/lox(ex3)) mice with Col1a1-cre mice. In mutant mice, tooth formation and eruption was disturbed. Lower incisors and molars did not erupt. Bone formation was increased in the mandible but tooth formation was severely disturbed. Hypomineralized dentin was deposited in the crown but roots of molars were extremely short and distorted. In the odontoblasts of mutant molars, expression of dentin matrix proteins was obviously downregulated following the activation of β-catenin whereas that of mineralization inhibitor was increased. Cementum and periodontal ligament were hypoplastic but periodontal space was narrow due to increased alveolar bone formation. While cementum matrix proteins were decreased, bone matrix proteins were increased in the cementum and alveolar bone of mutant mice. These results indicate that local activation of β-catenin in the osteoblasts and odontoblasts leads to aberrant dento-alveolar complex formation. Therefore, appropriate inhibition of Wnt/β-catenin signaling is important for the dento-alveolar complex formation.